Metabolic Pathway Design

Author : Chiam Ng
Publisher :
ISBN 13 :
Total Pages : pages
Book Rating : 4.:/5 (12 download)

DOWNLOAD NOW!

Download or read book Computational Design and Optimization of Metabolic Pathways written by Chiam Ng and published by . This book was released on 2017 with total page pages. Available in PDF, EPUB and Kindle. Book excerpt: Microbial production of chemicals and fuels has often been cited as the highly viable solution to the current energy crisis. It has also been widely exploited to produce value-added products such as pharmaceuticals, nutraceuticals, food flavorings and fine chemicals. Knowledge gained from genome sequencing and functional annotations, as well as advances in genetic engineering techniques, enable us to re-code the genetic blueprint of a microbial cell to endow them with new functions such as a non-native metabolic pathway that make a specific biochemical. As the cells are often not optimized to synthesize the target product, significant rewiring of their metabolic networks is required to re-apportion carbon flux towards the target product. This should be performed with careful consideration of cellular protein resource allocation as well as energy and redox balance. However, the task is often complicated by the intricate interplay of the cellular gene-protein-reaction network. In this dissertation, we aim to address several challenges in designing and engineering metabolic pathways by combining techniques in computational biology, metabolic engineering, and synthetic biology. Each project relies extensively on optimization-based computational design, from genetic constructs to pathways and finally the entire metabolic network, to achieve the associated metabolic engineering objectives. In Chapter 1, we review the state-of-the-art metabolic engineering and synthetic biology tools for pathway and strain design. These tools lay the foundation for the development of the subsequent chapters, which deal in-depth with specific challenges in metabolic pathway design and strain optimization. One of the key challenges in metabolic engineering is to ensure a consistent supply of cofactor, which is often shared across over 100 reactions in a cell, to the desirable biosynthesis reactions. In Chapter 2, we describe the design and optimization of a synthetic metabolic pathway in E. coli to improve the availability of an essential redox cofactor NADPH. The synthetic metabolic pathway was derived from the highly active Entner-Doudoroff (ED) pathway of another prokaryote Zymomonas mobilis. Upon optimizing the expression of the multi-enzyme pathway, we obtained strains with improved NADPH production when compared to the wild-type strain. In addition to NADPH production, the ED pathway also simultaneously generates pyruvate and glyceraldehyde-3-phopshate, which are the precursors for the methylerythritol phosphate (MEP) pathway. As a proof-of-concept, we demonstrated that combination of the synthetic ED pathway and the MEP pathway improved the production of an NADPH-dependent terpenoid by up to 97%. Motivated by the intriguing roles that glycolytic pathways play in a cell and the effects of their perturbation, in Chapter 3, we switch focus to uncover the reason why specific glycolytic pathways prevail in nature despite the presence of alternatives. Although the ED pathway and the Embden-Meyerhof-Parnas (EMP) glycolytic pathways both convert glucose to pyruvate, they take different routes with different intermediates and yield one or two moles of ATP per mole of glucose. Theoretically, one could construct a thermodynamically feasible (i.e., rG